JP2014504860A5

JP2014504860A5 -

Info

Publication number: JP2014504860A5
Application number: JP2013546325A
Authority: JP
Filing date: 2011-12-20
Publication date: 2015-02-05
Anticipated expiration: 2031-12-20

Claims

A binding protein comprising a polypeptide chain, wherein the polypeptide chain comprises VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first heavy chain variable domain;
VD2 is the second heavy chain variable domain;
C is a heavy chain constant domain;
X1 is a linker (but not CH1),
X2 is an Fc region,
n is 0 or 1. )
Including
A binding protein capable of binding to a pair of antigens selected from the group consisting of IL-1 alpha and IL-1 beta.

2. The binding protein of claim 1, wherein VD1 and VD2 comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 30, 32, 34, 36, 38, 40, 42, 44 and 46.

A binding protein comprising a polypeptide chain, wherein the polypeptide chain comprises VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first light heavy chain variable domain;
VD2 is the second light heavy chain variable domain;
C is a light chain constant domain;
X1 is a linker (but not CH1);
X2 does not contain an Fc region;
n is 0 or 1. )
Including
The binding proteins include IL-1 alpha (sequence 3) and IL-1 beta (sequence 1); IL-1 alpha (sequence 2) and IL-1 beta (sequence 1); IL-1 alpha (sequence 1) and IL-1 beta (sequence 1); IL-1 alpha (sequence 3) and IL-1 beta (sequence 2); IL-1 alpha (sequence 4) and IL-1 beta (sequence 2); IL-1 alpha ( Sequence 4) and IL-1 beta (sequence 3); consisting of IL-1 alpha (sequence 4) and IL-1 beta (sequence 4); IL-1 alpha (sequence 4) and IL-1 beta (sequence 5) A binding protein capable of binding to a pair of antigens selected from the group.

4. The binding protein of claim 3, wherein the VD1 and VD2 light chain variable domains comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 31, 33, 35, 37, 39, 41, 43, 45 and 47.

The binding protein according to claim 1 or 3, wherein n is 0.

A binding protein comprising first and second polypeptide chains, wherein the first polypeptide chain is a first VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first heavy chain variable domain;
VD2 is the second heavy chain variable domain;
C is a heavy chain constant domain;
X1 is a linker (but not CH1),
X2 is an Fc region. )
Wherein the second polypeptide chain comprises a second VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first light chain variable domain;
VD2 is the second light chain variable domain;
C is the light chain constant domain;
X1 is a linker (but not CH1),
X2 does not contain the Fc region,
n is 0 or 1. )
Including
The binding proteins include IL-1 alpha (sequence 3) and IL-1 beta (sequence 1), IL-1 alpha (sequence 2) and IL-1 beta (sequence 1), IL-1 alpha (sequence 1) and IL-1 beta (sequence 1), IL-1 alpha (sequence 3) and IL-1 beta (sequence 2), IL-1 alpha (sequence 4) and IL-1 beta (sequence 2), IL-1 alpha ( Sequence 4) and IL-1 beta (sequence 3), IL-1 alpha (sequence 4) and IL-1 beta (sequence 4), IL-1 alpha (sequence 4) and IL-1 beta (sequence 5) A binding protein capable of binding to a pair of antigens selected from the group.

The VD1 and VD2 heavy chain variable domains comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 30, 32, 34, 36, 38, 40, 42, 44 and 46, and the VD1 and VD2 light chain variable domains are sequences 7. A binding protein according to claim 6 comprising an amino acid sequence selected from the group consisting of numbers 31, 33, 35, 37, 39, 41, 43, 45 and 47.

The binding protein according to claim 1, 3 or 6, wherein X1 or X2 is an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 26.

7. The binding protein of claim 6, comprising two first polypeptide chains and two second polypeptide chains.

The binding protein according to claim 1, 3 or 6, wherein the Fc region is selected from the group consisting of a native sequence Fc region and a variant sequence Fc region.

11. The binding protein according to claim 10, wherein the Fc region is selected from the group consisting of IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgE and IgD derived Fc regions.

7. Binding according to claim 1, 3 or 6, wherein VD1 of the first polypeptide chain and VD1 of the second polypeptide chain are obtained from the same first and second parent antibody or antigen-binding portion thereof, respectively. protein.

The binding protein according to claim 1, 3 or 6, wherein VD1 of the first polypeptide chain and VD1 of the second polypeptide chain are obtained from different first and second parent antibodies or antigen-binding portions thereof, respectively. .

7. Binding according to claim 1, 3 or 6, wherein VD2 of the first polypeptide chain and VD2 of the second polypeptide chain are obtained from the same first and second parent antibodies or antigen-binding portions thereof, respectively. protein.

The binding protein according to claim 1, 3 or 6, wherein VD2 of the first polypeptide chain and VD2 of the second polypeptide chain are obtained from different first and second parent antibodies or antigen-binding portions thereof, respectively. .

16. A binding protein according to any one of claims 13 to 15 wherein the first and second parent antibodies bind to different epitopes on the antigen.

16. The first parent antibody or antigen binding portion thereof binds to the first antigen with a different potency than the second parent antibody or antigen binding portion thereof binds to the second antigen. The binding protein according to any one of the above.

14. The first parent antibody or antigen binding portion thereof binds to the first antigen with an affinity that is different from the affinity with which the second parent antibody or antigen binding portion thereof binds to the second antigen. To 15. The binding protein according to any one of 15 to 15.

The first parent antibody or antigen-binding portion thereof and the second parent antibody or antigen-binding portion thereof are selected from the group consisting of human antibodies, CDR-grafted antibodies and humanized antibodies. The binding protein according to claim 1.

A first parent antibody or antigen-binding portion thereof and a second parent antibody or antigen-binding portion thereof comprising a Fab fragment; an F (ab ′) ₂ fragment; and two Fab fragments joined by a disulfide bridge in the hinge region. Fd fragment consisting of VH and CH1 domains; Fv fragment consisting of VL and VH domains of an antibody single arm; dAb fragment; isolated complementarity determining region (CDR); single chain antibody; and diabody The binding protein according to any one of claims 13 to 15, which is selected from the group consisting of:

7. The binding protein of claim 1, 3 or 6 wherein the binding protein has at least one desired property exhibited by the first parent antibody or antigen binding portion thereof or the second parent antibody or antigen binding portion thereof.

23. A binding protein according to claim 22, wherein the desired property is selected from one or more antibody parameters.

Antibody parameters are antigen specificity, affinity for antigen, potency, biological function, epitope recognition, stability, solubility, production efficiency, immunogenicity, pharmacokinetics, bioavailability, tissue cross-reactivity and orthologous The binding protein according to claim 21, which is selected from the group consisting of antigen binding.

A binding protein capable of binding to two antigens, comprising four polypeptide chains, wherein the two polypeptide chains are VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first heavy chain variable domain;
VD2 is the second heavy chain variable domain;
C is a heavy chain constant domain;
X1 is a linker (but not CH1),
X2 is an Fc region. )
And the two polypeptide chains are VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first light chain variable domain;
VD2 is the second light chain variable domain;
C is the light chain constant domain;
X1 is a linker (but not CH1),
X2 does not contain the Fc region,
n is 0 or 1. )
Including
The VD1 and VD2 heavy chain variable domains comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 30, 32, 34, 36, 38, 40, 42, 44 and 46, and the VD1 and VD2 light chain variable domains are A binding protein comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 31, 33, 35, 37, 39, 41, 43, 45 and 47.

A binding protein capable of binding to two antigens, comprising four polypeptide chains, wherein the two polypeptide chains are VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first heavy chain variable domain;
VD2 is the second heavy chain variable domain;
C is a heavy chain constant domain;
X1 is a linker (but not CH1);
X2 is an Fc region;
n is 0 or 1. )
And the two polypeptide chains are VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first light chain variable domain;
VD2 is the second light chain variable domain;
C is a light chain constant domain;
X1 is a linker (but not CH1);
X2 does not contain an Fc region;
n is 0 or 1. )
Where DVD-Ig is IL-1 alpha (sequence 1); IL-1 alpha (sequence 2); IL-1 alpha (sequence 3); IL-1 alpha (sequence 4); At least selected from the group consisting of: beta (sequence 1); IL-1 beta (sequence 2); IL-1 beta (sequence 3); IL-1 beta (sequence 4); and IL-1 beta (sequence 5) A binding protein that binds to one antigen.

The binding protein is at least about 10 ² M ⁻¹ s ⁻¹ ; at least about 10 ³ M ⁻¹ s ⁻¹ ; at least about 10 ⁴ M ⁻¹ s ⁻¹ ; at least about 10 when measured by surface plasmon resonance. ^5. A binding rate constant (Kon) to one or more targets selected from the group consisting of: ⁵ M ⁻¹ s ⁻¹ ; and at least about 10 ⁶ M ⁻¹ s ⁻¹ . The binding protein according to 24 or 25.

The binding protein, as measured by surface plasmon resonance, is up to about 10 ⁻³ s ⁻¹ ; up to about 10 ⁻⁴ s ⁻¹ ; up to about 10 ⁻⁵ s ⁻¹ ; and up to about 10 ⁻⁶ s ^−1. 26. A binding protein according to claim 1, 3, 6, 24 or 25, having a dissociation rate constant (Koff) to one or more targets selected from the group consisting of.

Up to about 10 ⁻⁷ M; up to about 10 ⁻⁸ M; up to about 10 ⁻⁹ M; up to about 10 ⁻¹⁰ M; up to about 10 ⁻¹¹ M; up to about 10 ⁻¹² M for one or more targets 26. A binding protein according to claim 1, 3, 6, 24 or 25, having a dissociation constant (K _D ) selected from the group consisting of; and up to 10 ⁻¹³ M.

26. An action comprising the binding protein according to any one of claims 1, 3, 6, 24 or 25, wherein the binding protein conjugate is selected from the group consisting of immunoadhesion molecules, contrast agents, therapeutic agents and cytotoxic agents. A binding protein conjugate further comprising a substance.

30. The binding protein conjugate of claim 29, wherein the antigen is a contrast agent selected from the group consisting of a radioactive label, an enzyme, a fluorescent label, a luminescent label, a bioluminescent label, a magnetic label and biotin.

The contrast agent is a radioactive label selected from the group consisting of ³ H, ¹⁴ C, ³⁵ S, ⁹⁰ Y, ⁹⁹ Tc, ¹¹¹ In, ¹²⁵ I, ¹³¹ I, ¹⁷⁷ Lu, ¹⁶⁶ Ho and ¹⁵³ Sm. 31. A binding protein conjugate according to 30.

A therapeutic or cytotoxic agent wherein the agent is selected from the group consisting of metabolic inhibitors, alkylating agents, antibiotics, growth factors, cytokines, anti-angiogenic agents, anti-mitotic agents, anthracyclines, toxins and apoptotic agents 32. The binding protein conjugate of claim 30, wherein

26. A binding protein according to claim 1, 3, 6, 24 or 25, wherein the binding protein is a crystallized binding protein.

34. The binding protein according to claim 33, wherein the crystal is a pharmaceutical sustained release crystal containing no carrier.

34. The binding protein of claim 33, wherein the binding protein has a greater in vivo half-life than the soluble counterpart of the binding protein.

34. The binding protein of claim 33, wherein the binding protein retains biological activity.

26. An isolated nucleic acid encoding the binding protein amino acid sequence of any one of claims 1, 3, 6, 24 or 25.

38. A vector comprising the isolated nucleic acid of claim 37.

40. The vector of claim 38, wherein the vector is selected from the group consisting of pcDNA, pTT, pTT3, pEFBOS, pBV, pJV, pcDNA3.1 TOPO, pEF6TOPO and pBJ.

A host cell comprising the vector of claim 38.

41. The host cell according to claim 40, wherein the host cell is a prokaryotic cell.

The host cell is E. coli. 42. The host cell of claim 41, which is E. coli.

41. The host cell according to claim 40, wherein the host cell is a eukaryotic cell.

44. The host cell according to claim 43, wherein the eukaryotic cell is selected from the group consisting of protist cells, animal cells, plant cells and fungal cells.

44. The host cell according to claim 43, wherein the eukaryotic cell is an animal cell selected from the group consisting of a mammalian cell, an avian cell and an insect cell.

46. The host cell of claim 45, wherein the host cell is a CHO cell.

46. The host cell of claim 45, wherein the host cell is COS.

44. The host cell according to claim 43, wherein the host cell is a yeast cell.

49. The host cell according to claim 48, wherein the yeast cell is Saccharomyces cerevisiae.

46. The host cell according to claim 45, wherein the host cell is an insect Sf9 cell.

51. A method for producing a binding protein comprising culturing a host cell according to any one of claims 40 to 50 in a medium under conditions sufficient to produce the binding protein.

52. The method of claim 51, wherein 50% -75% of the produced binding protein is a bispecific tetravalent binding protein.

52. The method of claim 51, wherein 75% -90% of the produced binding protein is a bispecific tetravalent binding protein.

52. The method of claim 51, wherein 90% -95% of the produced binding protein is a bispecific tetravalent binding protein.

52. A protein produced according to the method of claim 51.

56. A pharmaceutical composition comprising the binding protein according to any one of claims 1-36 and 55 and a pharmaceutically acceptable carrier.

57. The pharmaceutical composition according to claim 56, further comprising at least one additional therapeutic agent.

Additional therapeutic agents are therapeutic agents, contrast agents, cytotoxic agents, angiogenesis inhibitors; kinase inhibitors; costimulatory molecule blockers; adhesion molecule blockers; anti-cytokine antibodies or functional fragments thereof; methotrexate; cyclosporine; FK506; detectable label or reporter; TNF antagonist; anti-rheumatic drug; muscle relaxant, anesthetic, nonsteroidal anti-inflammatory drug (NSAID), analgesic, anesthetic, sedative, local anesthetic, neuromuscular blocker, Antimicrobial agent, anti-psoriatic agent, corticosteroid, anabolic steroid, erythropoietin, immunization, immunoglobulin, immunosuppressant, growth hormone, hormone replacement agent, radiopharmaceutical, antidepressant, antipsychotic, stimulant, asthma drug , Beta agonists, steroids for inhalation, epinephrine or analogs, cytokines and It is selected from the group consisting of preparative Cain antagonists, pharmaceutical composition according to claim 57.

The pharmaceutical composition of claim 56 for treating a disease or disorder of the Target.

The disease is rheumatoid arthritis, osteoarthritis, juvenile chronic arthritis, septic arthritis, Lyme arthritis, psoriatic arthritis, reactive arthritis, spondyloarthritis, systemic lupus erythematosis, Crohn's disease, ulcerative colitis, inflammatory bowel Disease, insulin-dependent diabetes mellitus, thyroiditis, asthma, allergic disease, psoriasis, dermatitis, scleroderma, graft-versus-host disease, organ transplant rejection, organ transplant-related acute or chronic immune disease, sarcoidosis, atherosclerosis Arteriosclerosis, disseminated intravascular coagulation, Kawasaki disease, Graves' disease, nephrotic syndrome, chronic fatigue syndrome, Wegener's granulomatosis, Henoch-Schlein purpura, renal microscopic vasculitis, chronic active hepatitis, uveitis , Septic shock, toxic shock syndrome, septic syndrome, cachexia, infectious disease, parasitic disease, acquired immune deficiency syndrome, acute transverse spine Inflammation, Huntington's disease, Parkinson's disease, Alzheimer's disease, stroke, primary biliary cirrhosis, hemolytic anemia, malignancy, heart failure, myocardial infarction, Addison's disease, sporadic, hypoendocrine syndrome type I and multiple Endocrine hypofunction syndrome type II, Schmidt syndrome, adult (acute) respiratory distress syndrome, alopecia, alopecia areata, seronegative arthropathy, arthropathy, Reiter's disease, psoriatic arthropathy, ulcerative colon arthropathy, Enteropathic synovitis, Chlamydia, Yersinia and Salmonella-related arthropathy, spondyloarthropathy, atherosclerosis / atherosclerosis, atopic allergy, autoimmune bullous disease, pemphigus vulgaris, deciduous pemphigus , Pemphigoid, linear IgA disease, autoimmune hemolytic anemia, Coombs positive hemolytic anemia, acquired pernicious anemia, juvenile pernicious anemia, myalgic encephalomyelitis / b Jarfree disease, chronic mucocutaneous candidiasis, giant cell arteritis, primary sclerosis hepatitis, idiopathic autoimmune hepatitis, acquired immune deficiency syndrome, acquired immune deficiency related diseases, hepatitis B, hepatitis C, classification Impaired immunodeficiency (unclassifiable primary hypogammaglobulinemia), dilated cardiomyopathy, female infertility, ovarian dysfunction, early ovarian dysfunction, fibrotic lung disease, idiopathic interstitial pneumonia, post-inflammation Interstitial lung disease, interstitial pneumonia, connective tissue disease related interstitial lung disease, mixed connective tissue disease related lung disease, systemic sclerosis related interstitial lung disease, rheumatoid arthritis related interstitial lung disease, Systemic lupus erythematosus-related lung disease, dermatomyositis / polymyositis-related lung disease, Sjogren's disease-related lung disease, ankylosing spondylitis-related lung disease, vasculitic diffuse lung disease, hemoside Cis-related lung disease, drug-induced interstitial lung disease, fibrosis, radioactive fibrosis, obstructive bronchiolitis, chronic eosinophilic pneumonia, lymphocyte infiltrating lung disease, post-infectious interstitial lung disease , Ventilatory arthritis, autoimmune hepatitis, type 1 autoimmune hepatitis (classical autoimmune or lupoid hepatitis), type 2 autoimmune hepatitis (anti-LKM antibody hepatitis), autoimmune-mediated hypoglycemia, melanoma Type B insulin resistance, hypoparathyroidism, acute immune disease related to organ transplantation, chronic immune disease related to organ transplantation, osteoarthritis, primary sclerosing cholangitis, type 1 psoriasis, type 2 Psoriasis, idiopathic leukopenia, autoimmune neutropenia, kidney disease NOS, glomerulonephritis, renal microvasculitis, Lyme disease, discoid lupus erythematosus, idiopathic male infertility or NOS, Sperm autoimmunity, multiple sclerosis (all Subtype), sympathetic ophthalmitis, pulmonary hypertension secondary to connective tissue disease, Goodpasture syndrome, pulmonary symptoms of polyarteritis nodosa, acute rheumatic fever, rheumatoid spondylitis, Still's disease, systemic sclerosis, Sjogren's syndrome, Takayasu / arteritis, autoimmune thrombocytopenia, idiopathic thrombocytopenia, autoimmune thyroid disease, hyperthyroidism, goiter autoimmune hypothyroidism (Hashimoto's disease), atrophic self Immune hypothyroidism, primary myxedema, lens-induced uveitis, primary vasculitis, vitiligo acute liver disease, chronic liver disease, alcoholic cirrhosis, alcohol-induced liver injury, cholestasis (Choleocytosis), idiosyncratic liver disease, drug-induced hepatitis, non-alcoholic steatohepatitis, allergy and asthma, group B linkage Fungal (GBS) infections, mental disorders (eg, depression and schizophrenia), diseases mediated by Th2 and Th1 types, acute and chronic pain (various forms of pain), and lung, breast, gastric, Cancers such as bladder cancer, colon cancer, pancreatic cancer, ovarian cancer, prostate cancer and kidney cancer and hematopoietic malignancies (leukemia and lymphoma), abetalipoproteinemia, advanced cyanosis, acute and chronic parasitic or infectious processes Acute leukemia, acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), acute or chronic bacterial infection, acute pancreatitis, acute renal failure, adenocarcinoma, aerial ectopic pulsation, AIDS recognition Complex, alcohol-induced hepatitis, allergic conjunctivitis, allergic contact dermatitis, allergic rhinitis, allograft rejection, α-1-antitrypsin Deficiency, amyotrophic lateral sclerosis, anemia, angina, anterior horn cell degeneration, anti-cd3 treatment, antiphospholipid syndrome, anti-receptor hypersensitivity reactions, aortic and Aneurysm, aortic dissection, arterial hypertension, arteriosclerosis, arteriovenous fistula, ataxia, atrial fibrillation (persistent or paroxysmal), atrial flutter, atrioventricular block, B cell lymphoma, bone transplant rejection, bone marrow transplantation (BMT) rejection, leg block, Burkitt lymphoma, burn, cardiac arrhythmia, cardiac dysfunction syndrome, cardiac tumor, cardiomyopathy, cardiopulmonary bypass inflammatory response, cartilage transplant rejection, cerebellar cortical degeneration, cerebellar disease, disorder Or multifocal atrial tachycardia, chemotherapy-related disease, chronic myelogenous leukemia (CML), chronic alcohol , Chronic inflammatory lesions, chronic lymphocytic leukemia (CLL), chronic obstructive pulmonary disease (COPD), chronic salicylic acid poisoning, colorectal cancer, congestive heart failure, conjunctivitis, contact dermatitis, pulmonary heart, coronary artery disease, Creutzfeldt-Jakob disease, culture negative sepsis, cystic fibrosis, cytokine therapy related diseases, boxer dementia, demyelinating disease, dengue hemorrhagic fever, dermatitis, dermatological symptoms, diabetes, diabetes mellitus, diabetic arteriosclerosis Disease, diffuse Lewy body disease, dilated congestive cardiomyopathy, basal ganglia disease, middle-aged Down's syndrome, drug-induced motor disease induced by drugs that block central nerve dopamine receptors, drug sensitivity, Eczema, encephalomyelitis, endocarditis, endocrine disease, epiglottis, Epstein-Barr virus infection, erythema, extrapituitary and cerebellar disease, familial hemophagocytic lymphatic system Familial hematophagic lymphohistiocytosis, fatal thymus transplant rejection, Friedreich ataxia, functional peripheral arterial disease, myocardial sepsis, gas gangrene, gastric ulcer, glomerulonephritis, any organ or tissue graft Rejection, Gram-negative sepsis, Gram-positive sepsis, granulomas caused by intracellular organisms, hairy cell leukemia, Hallerholden-Spatz disease, Hashimoto thyroiditis, hay fever, heart transplant rejection, hemochromatosis, hemodialysis, hemolytic uremic Syndrome / thrombolytic thrombocytopenic purpura, bleeding, hepatitis (type A), His bundle arrhythmia, HIV infection / HIV neuropathy, Hodgkin's disease, hyperkinetic disease, hypersensitivity reaction, hypersensitivity pneumonia, hypertension, hypokinetic disease, thalamus Lower-pituitary-adrenocortical system evaluation, idiopathic Addison disease, idiopathic pulmonary fibrosis, antibody vehicle Cytotoxicity, asthenia, infant spinal muscular atrophy, aortic inflammation, influenza A, ionizing radiation exposure, iridocyclitis / uveitis / optic neuritis, ischemia reperfusion injury, ischemic stroke, juvenile Rheumatoid arthritis, juvenile spinal muscular atrophy, Kaposi's sarcoma, renal transplant rejection, Legionella, leishmaniasis, leprosy, corticospinal lesions, lipemia, liver transplant rejection, lymphedema, Malaria, malignant lymphoma, malignant histiocytosis, malignant melanoma, meningitis, meningococcusemia, metabolic / idiopathic, migraine, mitochondrial multisystem. system disorder), mixed connective tissue disease, monoclonal hypergammaglobulinemia, multiple myeloma, multiple lineage degeneration (Mensell Degeline-Thomas Shy-Drager and Machado-Joseph), myasthenia gravis, Mycobacterium Abium intracellulare, mycobacterial tubaculosis, myelodysplasia, myocardial infarction, myocardial ischemic disease, nasopharyngeal cancer, neonatal chronic lung disease, nephritis, nephrosis, neurodegenerative disease, neurogenic I muscle atrophy, Neutropenic fever, non-Hodgkin lymphoma, obstruction of the abdominal aorta and its branches, obstructive arterial disease, okt3 therapy, testitis / epididymis, testicularitis / vaginal reconstruction, organ hypertrophy, osteoporosis, Pancreatic transplant rejection, pancreatic cancer, paraneoplastic syndrome / malignant hypercalcemia, parathyroid transplant rejection, pelvic Inflammatory disease, perennial rhinitis, pericardial disease, peripheral atherosclerotic disease, peripheral vascular disease, peritonitis, pernicious anemia, Pneumocystis carinii pneumonia, pneumonia, POEMS syndrome (polyneuritis, organ hypertrophy, endocrine disease, monoclonal gamma globulinemia and skin change syndrome), post perfusion syndrome, post pump syndrome, post-myocardial infarction open heart surgery syndrome, preeclampsia, progressive supranuclear palsy, primary pulmonary hypertension , Radiotherapy, Raynaud's phenomenon and disease, Raynaud's disease, refsum disease, regular narrow QRS tachycardia, renovascular hypertension, reperfusion injury, restrictive cardiomyopathy, sarcoma, scleroderma Senile dementia, Lewy body senile dementia, serum reaction Negative arthritis, shock, sickle cell anemia, cutaneous allograft rejection, skin change syndrome, small intestine transplant rejection, solid tumor, intrinsic arrhythmia, spinal ataxia, spinocerebellar degeneration, streptococcal myositis, cerebellar structural lesions , Subacute sclerosing panencephalitis, eczema, cardiovascular syphilis, systemic anaphylaxis, systemic inflammatory response syndrome, systemic onset juvenile rheumatoid arthritis, T cells or FABALL, telangiectasia, obstructive thromboangiitis, Thrombocytopenia, toxicity, transplantation, trauma / bleeding, type III hypersensitivity reaction, type IV hypersensitivity, unstable angina, uremia, urinary sepsis, hives, valvular heart disease, varicose veins, vasculitis, vein Disease, venous thrombosis, ventricular fibrillation, viral and fungal infection, viral encephalitis / aseptic meningitis, virus-related hemophagocytic syndrome, Werni Ke-Korsakoff syndrome, Wilson disease, xenograft rejection of any organ or tissue, acute coronary syndrome, acute idiopathic polyneuropathy, acute inflammatory demyelinating polyradiculoneuropathy, acute ischemia, adult Still disease Alopecia areata, anaphylaxis, antiphospholipid antibody syndrome, aplastic anemia, arteriosclerosis, atopic eczema, atopic dermatitis, autoimmune dermatitis, autoimmune abnormalities associated with streptococcal infection, autoimmune bowel Disease, autoimmune hearing loss, autoimmune lymphoproliferative syndrome (ALPS), autoimmune myocarditis, autoimmune early ovarian failure, blepharitis, bronchiectasis, bullous pemphigoid, cardiovascular disease, fulminant type Antiphospholipid syndrome, celiac disease, cervical spondylosis, chronic ischemia, scarring pemphigus, clinical sporadic syndrome (cis) at risk for multiple sclerosis, conjunctivitis, childhood-onset psychiatric disorder, chronic Obstructive pulmonary disease (COPD), lacrimal cystitis, dermatomyositis, diabetic retinopathy, diabetes mellitus, herniated disc, prolapse of the disc, drug-induced immune hemolytic anemia, endocarditis, endometriosis, endophthalmitis, Superior scleritis, polymorphic erythema, severe polymorphic erythema, gestational pemphigoid, Guillain-Barre syndrome (GBS), hay fever, Hughes syndrome, idiopathic Parkinson's disease, idiopathic interstitial pneumonia, IgE Mediated allergy, immune hemolytic anemia, inclusion body myositis, infectious ocular inflammatory disease, inflammatory demyelinating disease, inflammatory heart disease, inflammatory kidney disease, IPF / UIP, iritis, keratitis, dry keratoconjunctivitis, Kusmaul's disease or Kusmaul-Meyer's disease, Laundry paralysis, Langerhans cell histiocytosis, reticulated macula, macular degeneration, microscopic polyangiitis, Morbus bechterev, exercise new Disease, mucous pemphigoid, multiple organ failure, myasthenia gravis, myelodysplastic syndrome, myocarditis, radiculopathy, neuropathy, non-A non-B hepatitis, optic neuritis, osteolysis, ovarian cancer, small Arthritic JRA, peripheral artery occlusion disease (PAOD), peripheral vascular disease (PVD), peripheral arterial disease (PAD),
Phlebitis, polyarteritis nodosa (or periarteritis nodosa), polychondritis, polymyalgia rheumatica, leukosis, polyarticular JRA, polyendocrine deficiency syndrome, polymyositis, rheumatic Multiple myalgia (PMR), postpump syndrome, primary parkinsonism, prostate and rectal cancer and hematopoietic malignancies (leukemia and lymphoma), prostatitis, erythroblastic fistula, primary adrenal insufficiency, recurrent optic nerve spinal cord Inflammation, restenosis, rheumatic heart disease, sapho (synovitis, acne, pustulosis, bone hyperplasia and osteomyelitis), scleroderma, secondary amyloidosis, shock lung, scleritis, sciatica, secondary Adrenal insufficiency, silicone-related connective tissue disease, Snedon-Wilkinson dermatosis, ankylosing spondylitis, Stevens-Johnson syndrome (SJS), systemic inflammatory response syndrome, temporal arteritis, toxopra Retinitis mellitus, toxic epidermis, transverse myelitis, TRAPS (tumor necrosis factor receptor), type 1 allergic reaction, type II diabetes, hives, normal interstitial pneumonia (UIP), vasculitis, spring 60. The pharmaceutical composition of claim 59, selected from the group comprising conjunctivitis, viral retinitis, Vogt-Koyanagi-Harada syndrome (VKH syndrome), wet macular degeneration, wound healing, Yersinia and Salmonella-related arthropathy.

Parenteral, subcutaneous, intramuscular, intravenous, intraarticular, intrabronchial, intraperitoneal, intracisternal, cartilage, intracavity (intracavity), intracavity (intracelial), the cerebellum, intraventricular, colon, the cervical, stomach Intra, intrahepatic, intramyocardial, intraosseous, intrapelvic, pericardial, intraperitoneal, intrapleural, intraprostatic, intrapulmonary, intracolonial, intrarenal, intraretinal, intrathecal, intrasynovial, intrathoracic, uterus 61. The pharmaceutical composition according to claim 60, wherein the composition is administered to the subject by at least one mode selected from internal, intravesical, bolus, vaginal, rectal, oral, sublingual, intranasal and transdermal.

a) obtaining a first parent antibody or antigen-binding portion thereof capable of binding to the first antigen;
b) obtaining a second parent antibody or antigen-binding portion thereof capable of binding to a second antigen;
c) VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first heavy chain variable domain obtained from the first parent antibody or antigen-binding portion thereof;
VD2 is a second heavy chain variable domain obtained from the second parent antibody or antigen-binding portion thereof,
C is a heavy chain constant domain;
X1 is a linker (but not CH1),
X2 is an Fc region,
n is 0 or 1. Building first and third polypeptide chains comprising:
d) VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first light chain variable domain obtained from the first parent antibody or antigen-binding portion thereof;
VD2 is a second light chain variable domain obtained from the second parent antibody or antigen-binding portion thereof,
C is the light chain constant domain;
X1 is a linker (but not CH1),
X2 does not contain the Fc region,
n is 0 or 1. And e) producing a dual variable domain immunoglobulin capable of binding to the first antigen and the second antigen. A method of making a dual variable domain immunoglobulin capable of binding two antigens comprising expressing a first, second, third and fourth polypeptide chain comprising the steps of:
The binding proteins include IL-1 alpha (sequence 3) and IL-1 beta (sequence 1), IL-1 alpha (sequence 2) and IL-1 beta (sequence 1), IL-1 alpha (sequence 1) and IL-1 beta (sequence 1), IL-1 alpha (sequence 3) and IL-1 beta (sequence 2), IL-1 alpha (sequence 4) and IL-1 beta (sequence 2), IL-1 alpha ( Sequence 4) and IL-1 beta (sequence 3), IL-1 alpha (sequence 4) and IL-1 beta (sequence 4), IL-1 alpha (sequence 4) and IL-1 beta (sequence 5) A method capable of binding to a pair of antigens selected from the group.

The VD1 and VD2 heavy chain variable domains comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 30, 32, 34, 36, 38, 40, 42, 44 and 46, and the VD1 and VD2 light chain variable domains are sequences 63. The method of claim 62, comprising an amino acid sequence selected from the group consisting of numbers 31, 33, 35, 37, 39, 41, 43, 45 and 47.

63. The method of claim 62, wherein the first parent antibody or antigen binding portion thereof and the second parent antibody or antigen binding portion thereof are selected from the group consisting of human antibodies, CDR-grafted antibodies and humanized antibodies. .

A first parent antibody or antigen-binding portion thereof and a second parent antibody or antigen-binding portion thereof comprising a Fab fragment, an F (ab ′) ₂ fragment, and two Fab fragments joined by a disulfide bridge in the hinge region. From a valence fragment, an Fd fragment consisting of VH and CH1 domains, an Fv fragment consisting of VL and VH domains of a single arm of an antibody, a dAb fragment, an isolated complementarity determining region (CDR), a single chain antibody and a diabody 64. The method of claim 62, wherein the method is selected from the group consisting of:

64. The method of claim 62, wherein the first parent antibody or antigen binding portion thereof has at least one desired property exhibited by a dual variable domain immunoglobulin.

64. The method of claim 62, wherein the second parent antibody or antigen binding portion thereof has at least one desired property exhibited by a dual variable domain immunoglobulin.

64. The method of claim 62, wherein the Fc region is selected from the group consisting of a native sequence Fc region and a variant sequence Fc region.

64. The method of claim 62, wherein the Fc region is selected from the group consisting of Fc regions from IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgE, and IgD.

68. The method of claim 66, wherein the desired property is selected from one or more antibody parameters.

68. The method of claim 67, wherein the desired property is selected from one or more antibody parameters.

Antibody parameters are antigen specificity, affinity for antigen, potency, biological function, epitope recognition, stability, solubility, production efficiency, immunogenicity, pharmacokinetics, bioavailability, tissue cross-reactivity and orthologous 72. The method of claim 70, selected from the group consisting of antigen binding.

Antibody parameters are antigen specificity, affinity for antigen, potency, biological function, epitope recognition, stability, solubility, production efficiency, immunogenicity, pharmacokinetics, bioavailability, tissue cross-reactivity and orthologous 72. The method of claim 71, selected from the group consisting of antigen binding.

63. The first parent antibody or antigen binding portion thereof binds to the first antigen with an affinity that is different from the affinity with which the second parent antibody or antigen binding portion thereof binds to the second antigen. The method described in 1.

64. The first parent antibody or antigen binding portion thereof binds to the first antigen with a potency that is different from the potency that the second parent antibody or antigen binding portion thereof binds to the second antigen. the method of.

a) obtaining a first parent antibody or antigen-binding portion thereof that can bind to a first antigen and have at least one desired property exhibited by a dual variable domain immunoglobulin;
b) obtaining a second parent antibody or antigen-binding portion thereof that can bind to a second antigen and can have at least one desired property exhibited by a dual variable domain immunoglobulin;
c) VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first heavy chain variable domain obtained from said first parent antibody or antigen binding portion thereof;
VD2 is a second heavy chain variable domain obtained from said second parent antibody or antigen binding portion thereof;
C is a heavy chain constant domain;
X1 is a linker (but not CH1);
X2 is an Fc region;
n is 0 or 1. )
Constructing first and third polypeptide chains comprising:
d) VD1- (X1) n-VD2-C- (X2) n
(VD1 is the first light chain variable domain obtained from the first parent antibody or antigen-binding portion thereof;
VD2 is a second light chain variable domain obtained from said second parent antibody or antigen binding portion thereof;
C is a light chain constant domain;
X1 is a linker (but not CH1);
X2 does not contain an Fc region;
n is 0 or 1. )
Constructing second and fourth polypeptide chains comprising:
e) the first, second, third and fourth polypeptides so that a double variable domain immunoglobulin capable of binding to the first and second antigens having the desired properties is produced. A method of making a dual variable domain immunoglobulin capable of binding two antigens having desired properties, comprising expressing a chain, wherein the binding protein comprises IL-1 alpha (sequence 3) and IL IL-1 alpha (sequence 1); IL-1 alpha (sequence 2) and IL-1 beta (sequence 1); IL-1 alpha (sequence 1) and IL-1 beta (sequence 1); IL-1 alpha (sequence) 3) and IL-1 beta (sequence 2); IL-1 alpha (sequence 4) and IL-1 beta (sequence 2); IL-1 alpha (sequence 4) and IL-1 beta (sequence 3); IL- 1 A method capable of binding to a pair of antigens selected from the group consisting of Rufa (sequence 4) and IL-1 beta (sequence 4); IL-1 alpha (sequence 4) and IL-1 beta (sequence 5) .